Olfactomedin 4 modulates neutrophil killing of S. aureus and E. coli in mice through inhibiting cathepsin C-mediated protease activities Neutrophils kill bacteria generally through oxidative and nonoxidative mechanisms. Whereas much research has focused on the enzymes essential for neutrophil killing, little is known about the regulatory molecules responsible for such killing. In this study we investigated the role of olfactomedin 4 (OLFM4), an olfactomedin-related glycoprotein, in neutrophil bactericidal capability and host innate immunity. Neutrophils from OLFM4-/- mice have increased intracellular killing of Staphylococcus aureus and Escherichia coli in vitro. The OLFM4-/- mice have enhanced in vivo bacterial clearance and are more resistant to sepsis when challenged with S. aureus or E. coli by intraperitoneal injection. OLFM4 was found to interact with cathepsin C, a cysteine protease that plays an important role in bacterial killing and immune regulation. We demonstrated that OLFM4 inhibited cathepsin C activity in vitro and in vivo. The cathepsin C activity in neutrophils from OLFM4-/- mice was significantly higher than that in neutrophils from wild-type littermate mice. The activities of three serine proteases (neutrophil elastase, cathepsin G, and proteinase 3), which require cathepsin C activity for processing and maturity, were also significantly higher in OLFM4-/- neutrophils. The bacterial killing and clearance capabilities observed in OLFM4-/- mice that was enhanced relative to WT mice was significantly compromised by the additional loss of cathepsin C in mice with OLFM4 and cathepsin C double deficiency. These results indicate that OLFM4 is an important negative regulator of neutrophil bactericidal activity by restricting cathepsin C activity and its downstream granule-associated serine proteases. Deletion of OLFM4 Rescues Defective Host Defense Against Staphylococcus aureus, but not Aspergillus fumigatus in Murine X-linked Chronic Granulomatous Disease Chronic granulomatous disease (CGD) patients have recurrent life-threating bacterial and fungal infections due to the mutation of one of four subunits of the respiratory burst oxidase (NADPH-oxidase). Olfactomedin 4 (OLFM4) is a neutrophil granule protein that negatively regulates host defense against bacteria infection. We aimed to evaluate the impact of OLFM4 deletion on host defense against Staphylococcus aureus and Aspergillus fumigatus in a murine X-linked CGD model. We found that intracellular killings as well as in vivo clearance of S. aureus, and resistance to S. aureus sepsis were significantly increased in gp91phox-and OLFM4-double deficient mice compared with CGD mice. However, double deficiency had no effect on host defense against pulmonary infection by A. fumigatus. These results show that OLFM4 deletion can successfully rescue immune deficiency against S. aureus, but not A. fumigatus in CGD mice. OLFM4 might prove to be an important target in CGD patients to augment host defense against bacterial infection.